Lovastatin, pravastatin, compactin, and derivatives and analogs thereof, are known to be potent HMG-CoA reductase inhibitors and are used as antihyper-cholesterolemic agents. Lovastatin, compactin and pravastatin are produced by fermentation, using microorganisms of different species belonging to Aspergillus, Penicilium and Streptomyces genera, respectively.
The purity of the active ingredient is an important factor for manufacturing a safe and effective pharmaceutical, especially if the pharmaceutical product must be taken on a long-term basis in the treatment or prevention of high plasma cholesterol. Accumulation of impurities from pharmaceuticals of lower purity may cause many side effects during medical treatment of a subject.
Among all the statins produced by microorganisms, pravastatin is has significant advantages in that it exhibits stronger and highly tissue-selective inhibition of cholesterol synthesis (Tsujita et. al, Biochim Biophy Acta, 1986, 877, 50–60). Pravastatin is produced by microbial hydroxylation of its precursor, Compactin (also called ML-236B). This bioconversion is carried out by a number of microorganisms e.g. Streptomyces (U.S. Pat. No. 5,179,013, U.S. Pat. No. 4,448,979), Nocardia, Amycolata, Saccharopolyspora, Amycolatopsis, Saccharothrix, Gilbertella (EP 0649907, WO 99/60151), Actinomadura (WO 96/40863), Mortierella (WO 00/46175), Nocardia (U.S. Pat. No. 5,830,695) and Bacillus sp. (U.S. Pat. No. 6,245,535, WO 99/07827). A number of species of Streptomyces, e.g. S. carbophilus, S. hastedii (JP 4,349,034), S. flavovirens (WO 99/10419), S. rosenchromogenous (U.S. Pat. No. 4,346,227), S. californicus (EP 649907) and S. exfoliatus (WO 98/45410) are also known to carry out this bioconversion.
The bioconversion is a cytochrome p450 dependent system and the enzymes are induced by the presence of compactin in the medium (Matsuoka et al, European Journal of Biochemistry, 1989: 184: 707–713; Serizawa et. al, In: Biotechnology of antibiotics; W R Strohl (editor) 1997). Compactin must often be added into the seed medium for efficient bioconversion (WO 98/45410). Attempts have been made to clone and express this system by recombinant DNA techniques in a fungal host (e.g. Penicillium citrinum) for one step de novo production of pravastatin (WO 99/10499). However, the yields are not economically viable. Bioconversion using Streptomyces species is still currently the most efficient method of pravastatin production.